TY - JOUR
T1 - An assay to image neuronal microtubule dynamics in mice
AU - Kleele, Tatjana
AU - Marinković, Petar
AU - Williams, Philip R.
AU - Stern, Sina
AU - Weigand, Emily E.
AU - Engerer, Peter
AU - Naumann, Ronald
AU - Hartmann, Jana
AU - Karl, Rosa M.
AU - Bradke, Frank
AU - Bishop, Derron
AU - Herms, Jochen
AU - Konnerth, Arthur
AU - Kerschensteiner, Martin
AU - Godinho, Leanne
AU - Misgeld, Thomas
N1 - Funding Information:
We would like to thank Manuela Budak, Ljiljana Marinković and Nebahat Budak for animal husbandry; Sarah Bechtold, Yvonne Hufnagel and Kristina Wullimann for technical assistance, Monika Schetterer for administrative support and Gabriela Plu-cińska for help with initially testing the EB3-YFP construct in zebrafish. We thank Michael Coleman (Babraham Institute) for the DNLS-WldS mice and Brian Link (Medical College of Wisconsin) for the EB3-GFP construct. We thank Ed Ruthazer for point out the CANDLE algorithm to us. T.M., P.M., J.H. and F.B. are supported by the German Center for Neurodegenerative Disease (DZNE). Further support came from the Deutsche Forschungsgemeinschaft (DFG) via the Center for Integrated Protein Science Munich (EXC 114; A.K., T.M.), the Munich Cluster for Systems Neurology (EXC 1010 SyNergy; J.H., A.K., M.K., T.M.), Transregio 128 (M.K.), Priority Programme 1710 (M.K., T.M.), Research Training Group 1373 (P.E.), Collaborative Research Center 870 (A.K., L.G., T.M.) and Sonderforschungsbereich 1191 (F.B.). M.K.’s laboratory is further financed by the German Federal Ministry of Research and Education (BMBF; Competence Network Multiple Sclerosis), the European Research Council under the European Union’s Seventh Framework Programme (FP/2007–2013; ERC Grant Agreement n. 310932), the Hertie-Foundation and the ‘Verein Therapieforschung für MS-Kranke e.V.’. T.M. is supported by the European Research Council under the European Union’s Seventh Framework Programme (FP/2007–2013; ERC Grant Agreement n. 616791). F. B. is also supported by the International Foundation for Research in Paraplegia and Wings for Life. Work on this project was further supported by the national funding agency (BMBF) in the frame of ERA-Net ‘2-photon imaging’ (A.K., T.M.). P.R.W. was supported by the Human Frontier Science Program and the Wings for Life Foundation. D.B.’s lab is supported by the National Science Foundation (Award #1126196). T.K. and P.M. were supported by the Graduate School of Technische Universität München (TUM-GS).
Publisher Copyright:
© 2014 Macmillan Publishers Limited. All rights reserved.
PY - 2014
Y1 - 2014
N2 - Microtubule dynamics in neurons play critical roles in physiology, injury and disease and determine microtubule orientation, the cell biological correlate of neurite polarization. Several microtubule binding proteins, including end-binding protein 3 (EB3), specifically bind to the growing plus tip of microtubules. In the past, fluorescently tagged end-binding proteins have revealed microtubule dynamics in vitro and in non-mammalian model organisms. Here, we devise an imaging assay based on transgenic mice expressing yellow fluorescent protein-tagged EB3 to study microtubules in intact mammalian neurites. Our approach allows measurement of microtubule dynamics in vivo and ex vivo in peripheral nervous system and central nervous system neurites under physiological conditions and after exposure to microtubule-modifying drugs. We find an increase in dynamic microtubules after injury and in neurodegenerative disease states, before axons show morphological indications of degeneration or regrowth. Thus increased microtubule dynamics might serve as a general indicator of neurite remodelling in health and disease.
AB - Microtubule dynamics in neurons play critical roles in physiology, injury and disease and determine microtubule orientation, the cell biological correlate of neurite polarization. Several microtubule binding proteins, including end-binding protein 3 (EB3), specifically bind to the growing plus tip of microtubules. In the past, fluorescently tagged end-binding proteins have revealed microtubule dynamics in vitro and in non-mammalian model organisms. Here, we devise an imaging assay based on transgenic mice expressing yellow fluorescent protein-tagged EB3 to study microtubules in intact mammalian neurites. Our approach allows measurement of microtubule dynamics in vivo and ex vivo in peripheral nervous system and central nervous system neurites under physiological conditions and after exposure to microtubule-modifying drugs. We find an increase in dynamic microtubules after injury and in neurodegenerative disease states, before axons show morphological indications of degeneration or regrowth. Thus increased microtubule dynamics might serve as a general indicator of neurite remodelling in health and disease.
UR - http://www.scopus.com/inward/record.url?scp=84908256346&partnerID=8YFLogxK
U2 - 10.1038/ncomms5827
DO - 10.1038/ncomms5827
M3 - Article
C2 - 25219969
AN - SCOPUS:84908256346
SN - 2041-1723
VL - 5
JO - Nature communications
JF - Nature communications
M1 - 4827
ER -